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S L Vonderfecht

Publications and source records attributed to S L Vonderfecht.

At least 37 records · Page 2Linked to original sources
PubMed1992

Human milk mucin inhibits rotavirus replication and prevents experimental gastroenteritis.

Acute gastrointestinal infections due to rotaviruses and other enteric pathogens are major causes of morbidity and mortality in infants and young children throughout the world. Breast-feeding can reduce the rate of serious gastroenteritis in infants; however, the degrees of protection offered against rotavirus infection vary in different populations. The mechanisms associated with milk-mediated protection against viral gastroenteritis have not been fully elucidated. We have isolated a macromolecular component of human milk that inhibits the replication of rotaviruses in tissue culture and prevents the development of gastroenteritis in an animal model system. Purification of the component indicates that the antiviral activity is associated with an acidic fraction (pI = 4.0-4.6), which is free of detectable immunoglobulins. Furthermore, high levels of antiviral activity are associated with an affinity-purified complex of human milk mucin. Deglycosylation of the mucin complex results in the loss of antiviral activity. Further purification indicated that rotavirus specifically binds to the milk mucin complex as well as to the 46-kD glycoprotein component of the complex. Binding to the 46-kD component was substantially reduced after chemical hydrolysis of sialic acid. We have documented that human milk mucin can bind to rotavirus and inhibit viral replication in vitro and in vivo. Variations in milk mucin glycoproteins may be associated with different levels of protection against infection with gastrointestinal pathogens.

Adult↗
PubMed1992

Diarrhea caused by a slow-growing Enterococcus-like agent in neonatal rats.

An enteropathogenic Enterococcus-like agent was isolated from a spontaneous outbreak of diarrhea that occurred in a colony containing neonatal rats. Diarrhea was experimentally reproduced in virus-antibody-free neonatal rats inoculated with this purified "enterococcus." Gram-positive cocci were adhered to the small intestinal villi of affected animals from which the organism was reisolated. The isolate's classification in the genus Enterococcus was confirmed by genetic probe; however, because of its unique fermentation pattern, it could not be definitively speciated. Indirect immunofluorescence assays indicate that this strain of enterococcus and Enterococcus hirae, another strain pathogenic for neonatal rats, differ antigenically. Enterococci should be considered as potential etiologic agents in outbreaks of diarrhea involving neonatal rats and future efforts directed to increasing our understanding of the pathophysiology of this disease.

Animals↗
PubMed1990

Growth of group A rotaviruses in a human liver cell line.

Recent observations in children with rotavirus gastroenteritis and in infant mice given rotavirus vaccine by oral administration suggest that this well-known gastrointestinal pathogen may infect the liver. To examine this possibility, the susceptibility of Hep G2 cells to infection with a variety of rotavirus strains was tested. These cells were used because they are considered to be well differentiated and exhibit many liver-specific functions. The Hep G2 cells supported the growth of the simian strain rhesus rotavirus (MMU 18006), a strain currently being used in vaccine trails, but did not support the growth of any human strain (D, DS1, Price or ST3). The rhesus rotavirus infection was cytopathic and resulted in release of lactate dehydrogenase. Rhesus rotavirus growth in Hep G2 cells displayed trypsin-enhanced infectivity and was inhibited by pretreatment of cells with Arthrobacter ureafaciens neuraminidase but not with neuraminidase from Clostridium perfringens. Hep G2 cells were also permissive for another simian strain (SA11), a bovine strain (UK) and single gene substitution reassortants containing VP7 (the major outer capsid neutralization protein) from a human rotavirus strain and the remaining 10 genes from either rhesus rotavirus or UK. In general, UK and its reassortants produced lower levels of antigen than did rhesus rotavirus and its reassortants. Hep G2 cells and other hepatic cell lines may prove to be useful tools to explore the hepatotropic potential of wild-type rotaviruses and candidate vaccine strains.

Antigens, Viral↗
PubMed1989

Monoclonal antibody assay for detection of double-stranded RNA and application for detection of group A and non-group A rotaviruses.

Fastidious viruses are generally detected in human body fluids by means of immunoassay or nucleic acid hybridization systems. These approaches can be difficult to apply to the detection of viruses which display variations in antigenic or genetic composition. Rotaviruses are examples of viruses which can display such variations. Recently identified antigenic variants, designated as non-group A rotaviruses, cannot be detected by immunoassays or nucleic acid hybridization assays which utilize reagents directed at group A rotavirus strains. The incomplete understanding of the extent of antigenic and genetic variation has inhibited the development of assay systems for all of the non-group A rotaviruses and has limited the study of their role in human disease. While rotaviruses display genetic variation, they all contain a genome which consists of double-stranded RNA. We utilized a monoclonal antibody to devise a sensitive assay for the measurement of double-stranded RNA and applied it to the detection of a wide range of rotaviruses. We found that the assay could detect double-stranded RNA from as few as 10 PFU of standard strains of group A rotaviruses. The assay system was also capable of detecting double-stranded RNA from several strains of group B rotaviruses isolated from calves, rats, and pigs at levels below those at which viral RNA could be visualized by means of polyacrylamide gel electrophoresis. When applied to the detection of double-stranded RNA in serial stools shed by rotavirus-infected children, the assay system was capable of detecting double-stranded RNA in samples in which antigen could not be detected by immunoassay. The specific nature of the double-stranded RNA detected by this assay system could be determined by the elution of the nucleic acids from the monoclonal antibody and the reaction of the RNA with specific nucleotide probes. The measurement of double-stranded RNA offers a potential method for the sensitive detection of a wide range of rotaviruses and other members of the family Reoviridae.

Animals↗
PubMed1989

Detection of group B rotavirus in fecal specimens by dot hybridization with a cloned cDNA probe.

Cloned cDNA copies were synthesized from the genomic RNA of the IDIR strain of group B rotavirus (GBR) isolated in Baltimore, Md. These clones were screened for hybridization with heterologous GBR to identify cDNA for use in dot hybridization experiments. In multiple screening experiments, cDNA clones derived from gene segment 3 provided the most intense hybridization signals. 32P-labeled probes were produced from one of the gene 3 clones, and these were employed in dot hybridization assays. Purified preparations of bovine GBR were detected in concentrations of greater than or equal to 0.5 ng, and GBR was detected in fecal specimens obtained from five of six infected calves. Four of six human fecal specimens containing the Baltimore strain of GBR were also positive in the hybridization assay, while GBR was identified in only one of the six specimens by means of immunoelectron microscopy. A fecal specimen obtained from a patient infected with the adult diarrhea rotavirus strain of GBR was also positive in the dot hybridization assay. Fecal specimens from uninfected humans, calves, and rats, as well as specimens containing group A rotaviruses, did not hybridize with the cloned cDNA probe.

Animals↗
PubMed1989

Pathology of infectious diarrhea of infant rats (IDIR) induced by an antigenically distinct rotavirus.

Suckling rats were inoculated with a group B rotavirus to determine the progression of the morphologic changes induced in the intestine by this virus. Several changes were observed by light microscopy 1 day after viral inoculation: shortening of small intestinal villi, villous epithelial necrosis, and villous epithelial syncytia. The lesions were most often present in the distal small intestine, although other small intestinal segments were affected to a lesser degree. By day 3 post-inoculation, epithelial necrosis, and syncytia were no longer present; however, the villous epithelium was disorganized and irregularly vacuolated, and intestinal crypt epithelium was hyperplastic. Alterations in villous height to crypt depth ratios were present in portions of the small intestine for the remainder of the 12-day study period. Epithelial syncytia appeared to form by the breakdown of the lateral interdigitating membranes of the absorptive villous epithelium. Viral particles, abundant in the syncytia, appeared to form from amorphous or reticular arrays of viral precursor material. Group B rotaviral antigens, as detected by indirect immunofluorescence, were present in large amounts in the small intestinal villous epithelium only on the first day after viral inoculation. These studies show that two important diagnostic features of group B rotaviral infections of rats, epithelial syncytia and viral antigen as determined by immunofluorescence, are present only on the first day of disease. These findings should be taken into consideration when attempting to diagnose disease induced by this agent.

Animals↗
PubMed1988

Kinetics of intestinal replication of group B rotavirus and relevance to diagnostic methods.

Non-group-A rotaviruses have been implicated with increasing frequency as causes of acute gastroenteritis in humans and other animals. However, the incidence and significance of infection with these agents, as well as appropriate diagnostic strategies for making these determinations, are largely unknown. Studies to make these determinations could be more accurately conducted if the relationship between the viral replication kinetics and the particular diagnostic method used is understood. We thus utilized the murine model of group B rotavirus infection to establish the viral replication kinetics by a variety of commonly used diagnostic methods. Enzyme immunoassay, routine negative-stain electron microscopy, solid-phase immunosorbent electron microscopy, polyacrylamide gel electrophoresis, and a dot hybridization assay were used in these studies. By enzyme immunoassay, 100% of experimentally infected suckling rats tested positive for group B rotaviral antigens at 1, 4, and 5 days postinoculation. However, only 70 and 20% of infected animals tested positive at days 2 and 3 postinoculation, respectively. Dot hybridization with a complementary DNA probe also suggested a biphasic pattern of viral antigen excretion. Evidence of the virus causing infectious diarrhea in infant rats was found only on day 1 postinoculation in samples examined by routine negative-stain electron microscopy and by polyacrylamide gel electrophoresis. Rotaviruslike particles were observed by solid-phase immunosorbent electron microscopy on days 1, 2, and 4 after viral inoculation suckling rats but were clearly the most numerous on day 1. Additionally, the enzyme immunoassay was used to quantitate the kinetics of group B rotaviral replication in the intestines of the experimentally infected animals. Levels of murine group B rotaviral antigens in intestinal samples peaked on days 1 and 4 postinoculation; however, only peak 1 represented actual intraepithelial replication of the virus. These studies thus indicate that early sample collection and selection of the appropriate diagnostic method are critical if the incidence and significance of group B and possibly other non-group-A rotaviral infections are to be accurately assessed.

Animals↗
PubMed1988

Enterococcus hirae implicated as a cause of diarrhea in suckling rats.

A Lancefield group D enteric streptococcus was isolated from diarrheic suckling rats that had been inoculated orally with stool from a diarrheic human. After oral administration of the organism to other suckling rats, diarrhea was reproduced, and the enteric streptococcus was reisolated. The brush border of small intestinal villi in affected animals was coated with numerous adherent gram-positive cocci. The organism was identified as Enterococcus hirae by a battery of biochemical tests. These and previous studies indicate that certain enterococci should be considered as etiologic agents of diarrheal disease in neonatal animals.

Animals↗
PubMed1988

Protease inhibitors suppress the in vitro and in vivo replication of rotavirus.

Rotaviruses are major causes of infectious gastroenteritis in humans and other animals. We found that a variety of protease inhibitors suppressed the replication of the SA-11 strain of rotavirus in MA-104 cell cultures. Three of these compounds, leupeptin, pentamidine, and bis (5-amidino-2-benzimidazolyl) methane (BABIM) also restricted the intestinal replication of the murine strain of rotavirus when protease inhibitor and virus were administered simultaneously to suckling mice. Repeated administration of BABIM resulted in significantly reduced levels of intestinal rotaviral antigen even if administration of the compound was begun as late as 48 h after viral inoculation. Additionally, BABIM-treated animals had significantly less intestinal replication of rotavirus than did placebo-treated controls when placed in a heavily rotavirus-contaminated environment. The use of protease inhibitors represents a novel approach to the control of this important gastrointestinal pathogen and is a potential modality for the prevention and treatment of diseases caused by other enteric viruses, for which proteolytic cleavage is necessary for efficient replication.

Animals↗
PubMed1988

Duration and pattern of asymptomatic rotavirus shedding by hospitalized children.

Fecal shedding of rotaviruses by infants and children without gastrointestinal symptoms has recently been recognized, but little is known about the duration or frequency of virus shedding in such patients. We therefore evaluated rotavirus shedding by 16 infants and children during the course of their prolonged hospitalizations. Repeated fecal specimens were obtained from these patients and examined for the presence of rotavirus by means of immunoassay, electron microscopy and polyacrylamide gel electrophoresis. Asymptomatic rotavirus shedding was observed in each of the 16 patients, and more than 1 episode of virus excretion was detected in 15 of these. Electropherotype analysis indicated that some of these episodes were probably the result of reinfection with different strains of rotavirus. Asymptomatic shedding may be important in transmission of rotaviruses. Its role should be taken into consideration in future investigations of the epidemiology and control of this nosocomial pathogen.

Diarrhea, Infantile↗
PubMed1987

Comparison of methods for detection of serum antibody to murine rotavirus.

Mice are frequently used as animal models for the study of rotaviral infections. Since natural infection is common in laboratory mice, it is important that rotaviral studies, as well as other studies utilizing suckling mice, employ animals of known immune status to murine rotavirus. A variety of homologous and heterologous enzyme immunoassay systems and an immunofluorescence technique were thus compared to determine the immunoassay that is most effective at detecting adult mice seropositive for rotaviral antibody. It was determined that a homologous enzyme immunoassay inhibition technique utilizing murine rotavirus-derived reagents was the most efficient serologic assay evaluated. A serologic response was consistently detected by this assay by 5 days after experimental rotaviral inoculation of adult mice. A homologous antibody-binding enzyme immunoassay, a heterologous inhibition enzyme immunoassay utilizing antigenically related simian rotavirus (SA-11) reagents, and an immunofluorescence technique utilizing Nebraska calf diarrhea virus antigens were found to be less sensitive for detecting serum antibody to murine rotavirus.

Animals↗
PubMed1986

Homotypic and heterotypic antibodies for prevention of experimental rotavirus gastroenteritis.

We investigated the abilities of homologous and heterologous antibody preparations to neutralize murine rotavirus in a mouse model of rotavirus infection. We found that incubation of virus with murine sera obtained from animals experimentally infected with the homologous epizootic diarrhea of infant mice strain of rotavirus resulted in inability of the virus to cause symptomatic disease in infant mice. Sera from nonimmune mice and mice infected or immunized with a heterotypic strain of rotavirus did not effectively neutralize virus in this system. On the other hand, neutralization was noted after incubation of virus with a number of immunoglobulins from other animal and human sources. The neutralizing activity of the preparations in the murine model correlated partially, but not completely, with the level of in vitro neutralizing antibody to the murine strain of rotavirus measured in a tissue culture system. In most cases, asymptomatic infection after feeding of the virus and the antibody preparation resulted in subsequent generation of an active immune response in infected animals.

Animals↗
PubMed1986

Identification of a bovine enteric syncytial virus as a nongroup A rotavirus.

An atypical or nongroup A rotavirus was identified in feces obtained from gnotobiotic calves in which fecal preparations originally derived during an epizootic of neonatal calf diarrhea had been serially passaged. The epizootic was previously reported to be caused by a noncharacterized viral agent that induced the formation of epithelial syncytia on small intestinal villi of experimentally infected calves. This bovine, nongroup A rotavirus was found to be antigenically related to a described atypical rotavirus of rats by immunofluorescence and by enzyme immunoassay. Complementary DNA derived from the atypical rat rotavirus cross hybridized with RNA obtained from the bovine virus, but not with RNA extracted from group A rotaviruses. Complementary DNA derived from SA-11 group A rotavirus cross hybridized with other group A rotavirus RNA, but not with RNA obtained from either the rat or bovine nongroup A isolates. Additionally, another similar, if not identical, bovine atypical rotavirus was identified in a second epizootic of neonatal calf diarrhea that occurred several hundred kilometers and 8 months apart from the original epizootic.

Animals↗
PubMed1985

Enzyme immunoassay inhibition assay for the detection of rat rotavirus-like agent in intestinal and fecal specimens obtained from diarrheic rats and humans.

An enzyme immunoassay inhibition assay was developed to detect rat rotavirus-like agent (RVLA) or antigenically related viruses in intestinal washings and homogenates obtained from diarrheic rats and in fecal specimens obtained from diarrheic humans. In this assay, RVLA antigens in a test sample inhibited the binding of a biotin-labeled anti-rat RVLA antibody preparation to rat RVLA antigens bound to the solid phase. Intestinal washings and homogenates obtained 1 day after RVLA infection of suckling rats inhibited the binding of the biotin-labeled antibody to the solid-phase rat RVLA antigens by 76 to 100%. The inhibition was blocked by RVLA immune rat serum but not by nonimmune rat serum. Of 27 fecal specimens obtained from diarrheic humans, 6 produced disease characteristic of rat RVLA infection when inoculated into suckling rats. Four of these six specimens produced greater than 50% inhibition in the enzyme immunoassay. Fecal specimens obtained from diarrheic humans that were determined to be negative for RVLA produced an average inhibition of 9.2%. This enzyme immunoassay appears to be a useful diagnostic and research tool for the study of infections with at least one of the antigenically distinct rotaviruses.

Animals↗
PubMed1984

Infectious diarrhea of infant rats produced by a rotavirus-like agent.

During the investigation of an outbreak of diarrhea in suckling rats, a virus morphologically identical to but antigenically distinct from rotaviruses was identified. The disease was characterized clinically by erythema and cracking and bleeding of the perianal skin associated with the excretion of poorly formed fecal pellets, liquid, and gas. Light microscopy-observable changes consisted of small intestinal villous atrophy, villous epithelial necrosis, and villous epithelial syncytial cell formation. The cytoplasm of the epithelial syncytial cells contained large numbers of 80-nm viral particles that were often associated with reticular aggregates of electron-dense material. Viral infection principally involved the luminal one-fourth to one-third of the intestinal villi as determined by indirect immunofluorescence. This rotavirus-like agent contained 11 double-stranded RNA segments; however, the migration pattern of these segments in polyacrylamide gels differed from the electrophoretic pattern which is characteristic of the typical rotaviruses. The agent had a buoyant density in CsCl of 1.36 to 1.4 g/cm3 and was labile at pH 3 and at 56 degrees C; however, infectivity of viral inocula was not altered by extensive treatment with ether or by pH 5 buffers. This disease, which we have named infectious diarrhea of infant rats, is the first recognized viral diarrhea of rats and appears to be a good model for the study of the recently recognized group of atypical rotaviruses.

Animals↗
PubMed1984

Pathology of larval Eustrongylides in the rabbit.

Three fishermen from Maryland who swallowed live bait-minnows developed severe abdominal pain within 24 hr; 2 required abdominal surgery. Larvae of the nematode Eustrongylides sp. were found in the peritoneal cavity of both (Guerin et al., 1982). In the current study, the lesions produced by Eustrongylides larvae were investigated in New Zealand white rabbits. None of these exhibited any signs of clinical illness; however, postmortem examination within 24 hr of inoculation revealed that larvae had migrated through the walls of the esophagus and stomach and viable larvae were recovered from the pleural and peritoneal cavities as well as from gastric contents. Necropsies performed at different intervals of time postinoculation showed that the migrating larvae had produced multi-focal peritonitis and multiple granulomata in the liver.

Animals↗